biochemistry3sf3fandomcom-20200213-history
Biochemistry3sf3 Wiki
Biochemistry 3SF3 Welcome to Biochemistry 3SF3: Biochemistry of Food Science Authors: '''Anish Nanda, Chen Yu Tang, Christopher Griffiths, Marc-Anthony Pinizzotto, Mohammed Mohiuddin, Nicholas Singh-Pickersgill, Raies Ahmed, Shannon Kainula Introduction Known for its various health benefits and nutrional value, yogurt is believed to be one of the oldest processed foods in human history. This creamy dairy product has long been incorporated into the staple diets of many cultures around the world. But how is it produced? What is the general process? What organisms are involved in the fermentation process? The purpose of this learning module is to briefly introduce the reader to the manufacturing process of yogurt and the importance and relevance of energy metabolism in the lactic acid bacteria to yogurt development. The Production of Yogurt '''Overview Yogurt is made by fermenting milk with bacteria such as Lactobacillus bulgaricus and Streptococcus thermophilus, giving yogurt its distinct flavor and texture.1 During fermentation, lactose in the milk is broken down into glucose, which is converted into pyruvate and then acetyl-coenzyme A under aerobic conditions. However, under anaerobic conditions, the pyruvate molecules cannot be converted into acetyl-coenzyme A. Instead, via catalysis by lactate dehydrogenase, pyruvate reacts with NADH to produce lactic acid.2 The lactic acid produced by the bacteria decreases the pH of yogurt, giving yogurt its sour taste. The bacteria also break down casein, the main protein in milk, giving yogurt its texture.3 Manufacturing Process3 The first step in yogurt manufacturing is the standardization of milk, in which milk is mixed with skim milk and cream to adjust the fat content of the product. At this point, stabilizing agents such as gelatin is also added to the milk to maintain the physical properties of yogurt, including its texture and appearance. Next, milk is homogenized: fat globules are disrupted into smaller globules, leading to an increase in surface area. The purpose of this step is to prevent fat separation later on in the manufacturing process, and to enhance the consistency of the yogurt. Milk is then treated with heat to kill any unwanted microorganisms. Following this, starter culture is then a dded to begin bacterial fermentation - convering lactose into lactic acid. The last step in this process is the cooling of the product to reduce further acid development.3 Fermentation Organisms The most pivotal step in the formation of yogurt from milk is fermentation. Following pasteurization, milk is cooled to 40-45°C and inoculated with the gram-positive thermophilic bacteria Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus.3 Both S. thermophilus and Lb. bulgaricus are bacteria capable of fermenting lactic acid in anaerobic conditions that thrive at the incubation temperature and pH of the milk.4 As these bacteria ferment lactic acid from lactose, the milk acidifies from pH 6.7 to below 4.6. The milk coagulates at a pH of about 5.2 to 5.4 and then is used to form either set or stirred yogurt.3 Specifically, the bacteria Lactobacillus bulgaricus and Streptococcus thermophilus can be characterized as facultative anaerobes.5,6 These bacteria are able to produce Adenosine Triphosphate (ATP) in the presence of oxygen, through aerobic respiration. However in the absence of oxygen, they can use fermentation in order to produce ATP. Metabolic Reactions Glycolysis is a fundamental process in the production of yogurt by thermophilic lactic acid bacteria. Two pyruvate molecules are obtained from the glycolysis of one glucose or one galactose molecule, and are further processed to obtain ATP - which is used in energy-requiring processes. In anaerobic conditions, lactic acid is generated as a by-product. The initial substrate for glycosis in lactic acid bacteria is primarily lactose, a carbohydrate present in milk. Lactose is taken up by different transporters in the two bacteria and then metabolized to obtain glucose and galactose.8 This reaction is catalyzed by the phospho-β-galactosidase enzyme in Lb. bulgaricus and by β-galactosidase in S. thermophilus.8 Galactose is then converted into glucose-6-phosphate through the Leloir pathway. Glucose-6-phosphate is also an intermediate in the metabolism of glucose. After pyruvate is formed via glycolysis, it is converted to lactic acid in what is known as anaerobic fermentation (anaerobic means in the absence of oxygen). Lactic acid is responsible for the acidification of milk. This results in the reduction of pH in the milk which plays a role in the denaturation of casein. This is important for the formation of yogurt and contributes to its structure and its flavor.8 Pyruvate is also the required substrate for aerobic metabolism in these bacteria. As mentioned previously, lactic acid bacteria are facultative anaerobes, meaning they can tolerate the presence of oxygen, but do not require it. They can use pyruvate to produce ATP via aerobic metabolism, however, this pathway yields hydrogen peroxide as a byproduct, which is toxic to them. Hydrogen peroxide effectively hinders their growth in higher concentrations, as they do not possess the enzyme catalase, responsible for breaking this byproduct down to water and oxygen.8 Therefore during yogurt production, milk is heat-treated prior to the addition of the bacteria in order to remove the dissolved oxygen.3 Symbiosis The independent effects of S. thermophilus and Lb. bulgaricus are not sufficient to form yogurt; their symbiotic interaction, known as proto-cooperation, is essential for the creation of yogurt.4 Their extensive proto-cooperation involves the production of formic acid, pyruvic acid, and carbon dioxide by S. thermophilus to accelerate the growth of Lb. bulgaricus, which in turn releases peptides and amino acids by to aid S. thermophilus.4 This interaction between species is largely responsible for the taste, smell, and structure of yogurt.4 While S. thermophilus and Lb. bulgaricus are certainly the most notable contributors to fermentation, they are often used in conjunction with other lactic acid bacteria which contribute to taste and other properties. These additional bacteria include Lb. casei, Lb. rhamnosus and other species from genus Bififdobacterium.4 Properties of Yogurt Conditions The optimum temperature for S. thermophilus and Lb. bulgaricus to perform their function in converting lactose to lactic acid, is between 40-45°C.3 The optimal pH for this process is at a low pH range: between 4.6-6.7. In the production of yogurt, these specific conditions have tremendous effects on the quality of the final product. In terms of the temperature used for incubation, yogurt that has been incubated at a lower temperature - corresponding to that of 40-45°C - is found to have a stronger network of linked gels and proteins, specifically casein proteins. These casein protein molecules are able to bind more effectively to other protein molecules, both due to the arrangement of the proteins and the increased surface area between particles.3 As a result, the yogurt has a much more firm and stiff composition, which is desirable. Additionally, yogurt that has been incubated within this temperature range is more viscous, well-coated, and smoother. Similarly, the pH of the yogurt as production is taking place has a significant effect on the interaction between casein protein molecules, which directly affects the physical properties of the yogurt. During production, yogurt is typically at a pH near 7.0 before acidification takes place.3 When the pH drops from 6.7 to approximately 4.6, the isoelectric point of casein is reached. As the pH approaches this isoelectric point, the negative charges present on casein molecules at higher pH are reduced. This in turn decreases the amount of electrostatic repulsion between casein molecules and thereby causes an increase in the electrostatic attraction. Hydrophobic interactions are also increased near the isoelectric point, contributing to stronger attraction between molecules. These effects result in the formation of strong three-dimensional network of chains and clusters of casein proteins, adding to the richness in texture and quality of the final product. Lactic Acid Lactic acid is the major product of milk fermentation, which is the central process of yogurt production. During yogurt production, fermentation is allowed to continue until a desired acidity is reached, which requires that a certain concentration of lactic acid be achieved, as it is the primary determinant of the yogurt’s acidity. As mentioned above, when the pH decreases, the yogurt gel begins to form due to the formation of a three-dimensional network of casein proteins. Lactic acid concentration therefore heavily impacts the texture of the yogurt, although other processing steps and modifications are usually involved for further improvement. For example, stirred yogurt is pumped through a screen to give the yogurt a smooth and viscous texture.3 The sour aftertaste of yogurt is attributable to lactic acid, while firmness and texture will depend on variables such as the temperature at which fermentation occurs and total solids content.1,3 Sensory Evaluation Consumer acceptance of a product is highly based upon sensory perception. Consumers prefer thicker, firmer yogurts with low whey separation and smooth texture, and a product will not do well on the market if it does not possess these qualities.3 It therefore behooves companies to conduct sensory evaluations in order to test their products’ popularity prior to sales. In addition, sensory evaluation becomes important where standard measurement methods do not exist for a specific physical property. For example, there is no set method for evaluating smoothness of texture.1 The Link to Diabetes Diabetes Mellitus Type 2 Diabetes mellitus type 2 is a metabolic disorder characterized by peripheral insulin resistance.9 Insulin is a hormone responsible for the absorption of glucose by the cells in the liver, skeletal muscle and fat tissue. As a result of insulin resistance, cells in the body are unable to properly absorb glucose from the bloodstream, leading to hyperglycemia and other health effects such as increased thirst, polyuria, fatigue, blurring of vision and weight loss.10 Specifically, resistance in muscle and fat cells significantly reduces glucose uptake, raising the blood glucose levels outside the normal range. In the presence of insulin, the liver decreases its secretion of glucose and increases glycogen synthesis, allowing for the storage of excess glucose in the body. However, insulin-resistant liver cells may fail to reduce glucose production and instead lower the rate of glycogen synthesis - ultimately contributing to an increase in blood glucose. 9 In addition, insulin resistance is also associated with elevated levels of free fatty acids in the plasma, as insulin resistance in adipose cells causes an increase in hydrolysis of stored triglycerides and reduced absorption of lipids from the bloodstream. The increase in plasma free fatty acid conentration induces an increase in hepatic glucose production, further increasing blood glucose levels.9 Elevated plasma free fatty acid levels are also believed to be responsible for insulin resistance itself.11 In order to compensate for the increase in glucose in the blood, the pancreas must continue to secrete more insulin to overcome the resistance. When an adequate amount of insulin cannot be produced, the blood glucose levels begin to rise and result in the individual developing type 2 diabetes.9 Increased consumption of dairy products is believed to be associated with a low incidence of type 2 diabetes and reduction in insulin resistance.12,13 Recently, one group of researchers found that consuming high amounts of low-fat dairy products may improve insulin resistance in affected individuals.14 Experimental Design 14 In this study, scientists randomly assigned participants into one of two treatment groups for a total of 6 months. The first group was a high dairy (HD) supplemented group in which each subject was instructed to consume 4 servings of dairy per day and the second group was a low dairy (LD) supplemented group, limited to no more than 2 servings of dairy per day. At the end of the initial 6-month period, the participants in the HD group were switched to the LD group for 6 months and vice versa. Subjects in the HD group were regularly provided with low fat dairy products for dietary incorporation, and were instructed to only consume the dairy products provided by the researchers. In addition, all individuals were required to maintain their normal diet (with the exception of their intake) and level of physical activity for the duration of the study. Although they initially began with 39 participants, only 23 managed to successfully complete the study. At the beginning, middle and end of each 6-month study phase, researchers measured each participant's metabolic responses and analyzed their blood plasma, body composition, energy expenditure and other factors associated with insulin resistance. Subjects were also asked to keep a record of their diet and and physical activity in a logbook. Results of the Study 14 Compared to their low-dairy counterparts, it was found that daily consumption of 4 servings of low-fat milk and yogurt by the high-dairy subjects reduced plasma insulin levels by an average of 9% and improved insulin resistance by 11% over 6 months. The authors speculate that these changes could be caused by several bioactive compounds present in dairy products, such as calcium and vitamin D. Vitamin D is able to sensitize insulin responses by regulating the expression of insulin receptors in cells, and by stimulating insulin release by pancreatic beta cells.15 However, no differences in vitamin D and calcium intake were observed. This may be due to poor dietary recall by subjects; since this was a "free-living" experiment (i.e. all subjects maintained their normal lifestyles throughout the study), it is prone to design limitations including the possibility of inaccurate diet records. Therefore, the authors were unable to identify specific bioactive components that may be responsible for the observed changes. In addition to the improvement in insulin resistance, no negative consequences were observed as a result of increased consumption of low-dairy products. Other measurements taken during the study - including blood pressure, blood glucose, and blood lipid and lipoprotein responses - remained consistent. The body weight, body composition and energy expenditure of participants in the high-dairy and low-dairy groups showed no significant changes either. These results are important as they suggest that the insulin-sensitizing effects of low-fat dairy products are not related to factors involved in energy metabolism. Long-term follow-up research could be performed in the future to verify the results from this study and possibly assay specific bioactive dairy-derived components to determine their effects on insulin resistance. It is clear, however, that consumption of dairy products is associated with insulin resistance, and research into this field could potentially lead to a novel treatment method for type 2 diabetes. Evaluation Think you've mastered the art and science of yogurt production? Test yourself! Evaluation: Solution Set (don't cheat!): Work through the evaluation first, and then check your answers using the solutions provided. The solution set contains the answer to every question, as well as appropriate feedback to go along. Having problems downloading the files? Try these links instead: Evaluation.pdf Solution_Set.pdf Works Cited 1. Horiuchi, H.; Inoue, N.; Liu, E.; Fukui, M.; Sasaki, Y.; Sasaki, T. A method for manufacturing superior set yogurt under reduced oxygen conditions. J. Dairy Sci. 2009, 92, 4112-4121. 2. Wee, Y. J.; Kim, J. N.; Ryu, H. W. Biotechnological production of lactic acid and its recent applications. Food Technology and Biotechnology 2006, 44, 163-172. 3. Lee, W. J.; Lucey, J. A. Formation and physical properties of yogurt. Asian-Aust. J. Anim. Sci 2010, 23, 1127-1136. 4. Angelov, M.; Kostov, G.; Simova, E.; Beshkova, D.; Koprinkova-Hristova, P. Proto-cooperation factors in yogurt starter cultures. e-Revue de Génie Industriel 2009, 3, 4-12. 5. Burgos-Rubio, C. N.; Okos, M. R.; Wankat, P. C. Kinetic study of the conversion of different substrates to lactic acid using Lactobacillus bulgaricus. Biotechnol. Prog. 2000, 16, 305-314. 6. Roussel, Y.; Pebay, M.; Guedon, G.; Simonet, J. M.; Decaris, B. Physical and genetic map of Streptococcus thermophilus A054. J. Bacteriol. 1994, 176, 7413-7422. 7. PSmicrographs. Yoghurt Bacteria Science Image. http://www.psmicrographs.co.uk/yoghurt-bacteria/science-image/80014272a (accessed October 16, 2013). 8. Zourari, A.; Accolas, J. P.; Desmazeaud, M. J. Metabolism and biochemical characteristics of yogurt bacteria. A review. Le lait 1992, 72, 1-34. 9. Reaven, G. M. Role of insulin resistance in human disease. Diabetes 1988, 37, 1595-1607. 10. Alberti, K. G.; Zimmet, P. Z. Definition, diagnosis and classification of diabetes mellitus and its complications. Part 1: diagnosis and classification of diabetes mellitus provisional report of a WHO consultation. Diabet. Med. 1998, 15, 539-553. 11. Itani, S. I.; Ruderman, N. B.; Schmieder, F.; Boden, G. Lipid-induced insulin resistance in human muscle is associated with changes in diacylglycerol, protein kinase C, and IkBeta-alpha. Diabetes 2002, 51, 2005-2011. 12. Stancliffe, R. A.; Thorpe, T.; Zemel, M. B. Dairy attentuates oxidative and inflammatory stress in metabolic syndrome. Am. J. Clin. Nutr. 2011, 94, 422-430. 13. Tong, X.; Dong, J. Y.; Wu, Z. W.; Li, W.; Qin, L. Q. Dairy consumption and risk of type 2 diabetes mellitus: a meta-analysis of cohort studies. Eur. J. Clin. Nutr. 2011, 65, 1027-1031. 14. Rideout, T. C.; Marinangeli, C. P.; Martin, H.; Browne, R. W.; Rempel, C. B. Consumption of low-fat dairy foods for 6 months improves insulin resistance without adversely affecting lipids or bodyweight in healthy adults: a randomized free-living cross-over study. Nutr. J. 2013, 12, 56-2891-12-56. 15: Borissova, A. M.; Tankova, T.; Kirilov, G.; Dakovska, L.; Kovacheva, R. The effect of vitamin D3 on insulin secretion and peripheral insulin sensitivity in type 2 diabetic patients. Int. J. Clin. Pract. 2003, 57, 258-261. Photos and videos are a great way to add visuals to your wiki. Find videos about your topic by exploring Wikia's Video Library. Category:Browse